Current methods are manual for detection of microorganisms. In such manual systems, a sample of material to be tested is incubated, usually in a suitable growth medium. Various manipulations such as agitation are required during the incubation and monitoring period. The detection of growth is achieved by visual inspection. For example, technicians observe and assess the growth of bacteria on a Petri dish, or evaluate the clarity of a broth (turbidity). The visual observations and assessments are subjective and, therefore, subject to error. In addition, these manual methods are labor intensive, require significant manipulation, and entail observation of all samples by laboratory personnel.
A number of methods have been suggested to detect the presence or absence of microorganisms by less subjective means. U.S. Pat. No. 3,743,581 (1973), Cady et al., discloses a method for monitoring microbiological growth by measuring the change in the conductivity of selected nutrient media inoculated with a sample.
U.S. Pat. No. 3,907,646 (1975), Wilkins et al, describes measurement of gas production of microorganisms. A pressure transducer is applied to a test tube and connected to a power source and strip recorders. Measurements are recorded on the strip recorders producing a plot of an electrical signal, which is generated over time, indicative of the presence and quantity of microorganisms. The instrument is very large and cumbersome, making it impractical to monitor multiple samples.
U.S. Pat. No. 4,152,213 describes a system by which the growth of microorganisms in a sealed container is detected by measuring reduction in headspace pressure as the microorganism consumes oxygen and comparing the reduction in pressure to a reference standard of the initial pressure. A vacuum sensor senses a reduction in pressure in the headspace of a container and provides an electrical signal to remote electronics. A major problem with such a system is that it is limited to those organisms that consume oxygen. Many microorganisms do not consume oxygen. Thus, the presence of a vacuum is not a universal indicator of microbial growth. Another problem with such a system is that in many instances the maximum decrease in the headspace pressure is small in comparison to the natural variations of the atmospheric pressure. In addition, this method requires precise pressure sensors since it functions on the basis of absolute value of initial and threshold pressures.
U.S. Pat. No. 5,232,839 describes a system by which the presence of microbiological growth in a sealed sample container is detected by measuring the rate of change of headspace pressure in the container as the microorganism consumes oxygen and comparing the change in pressure to a reference standard of the initial pressure. A vacuum sensor senses a reduction in pressure in the headspace of a container and provides an electrical signal to remote electronics. A major problem exists for weak consumers, or slower growing organisms, where background redox reactions can occur because the reagents added to the culture broth cause an unpredictable change in the pressure differential in the headspace due to reduction oxidation. A major problem with such a system is that it suffers from false positives due to background redox reactions.
It is well known that pH buffers are utilized for end point growth determinations using pH dyes, such as phenol red. The pH buffers are known to stabilize background pH drift due to chemical reactions within the test system. It is with this concept that one uses Redox buffer/dye systems to stabilize background chemical redox reactions, which can be applied generally to a detection system for microbial growth.
Use of poising agents to stabilize redox dyes for determination of end point growth reactions, such as antibiotic susceptibility has been described in U.S. Pat. No. 5,501,959 by Lancaster et al.
U.S. Pat. No. 6,395,506 discloses a device for monitoring cells for detection and evaluation of metabolic activity of eukaryotic and/or prokaryotic cells based upon their ability to consume dissolved oxygen. The methods utilize a luminescence detection system which makes use of the sensitivity of the luminescent emission of certain compounds to the presence of oxygen, which quenches (diminishes) the compound's luminescent emission in a concentration dependent manner.